Post-transcriptional regulation of neural polypyrimidine tract binding protein reprograms alternative splicing in neural and muscular cells

Neural PTB (nPTB/brPTB/PTBP2) is a closely related homolog of the splicing regulator polypyrimidine tract binding protein (PTB/hnRNP I/PTBPI). nPTB is largely restricted to neurons and testis, although it can be detected at lower levels in heart, skeletal muscle, and liver. nPTB protein is present in undifferentiated mouse myoblasts prior to their terminal differentiation into myotubes. During differentiation of C2C12 myoblasts, nPTB mRNA expression increases while the amount of nPTB protein in the cells is strongly reduced. We show that the muscle-restricted microRNA miR-133 represses nPTB protein expression without decreasing nPTB mRNA. A luciferase reporter linked to the nPTB 3' UTR is repressed by miR-133 through two miR-133 responsive elements (MRE). Both the miR-133 and coexpressed miR-1/206 microRNAs are conserved across animal species; we find that the PTB proteins contain conserved MREs for these RNAs in Drosophila, mice, and humans. The increase in miR-133 and the decrease in nPTB coincide with the induction of several alternatively spliced exons. Transfection of anti-sense oligonucleotides blocks the function of miR-133, increasing expression of nPTB and decreasing the inclusion of these exons. These results implicate microRNAs in the control of alternative splicing during development.; Studies of nPTB expression indicate that it is present in all major regions of the brain. We find that at the cellular level, nPTB protein is specifically expressed in postmitotic neurons whereas PTB is restricted to glia and other non-neuronal cells. nPTB protein expression is predominantly regulated at the post-transcriptional level. Depletion of PTB leads to efficient expression of nPTB. PTB represses nPTB expression through PTB-dependent alternative splicing of nPTB pre-mRNA leading to its destruction by nonsense-mediated decay (NMD). PTB also has a strong effect on nPTB translation or other processes independent of splicing. Our data suggest that the loss of PTB expression in developing neurons leads to increased expression of nPTB in mature neurons. PTB and nPTB show distinct patterns of splicing regulation of neuron-specific exons. In cell lines undergoing neuronal differentiation, when PTB levels decrease, nPTB is upregulated, suggesting that the changes in alternative splicing patterns of developing neurons are influenced by the switch from PTB to nPTB expression.